中国危重病急救医学
中國危重病急救醫學
중국위중병급구의학
CHINESE CRITICAL CARE MEDICINE
2011年
5期
275-278
,共4页
伊桐凝%张锦%于世家%高天舒
伊桐凝%張錦%于世傢%高天舒
이동응%장금%우세가%고천서
高糖%衰老细胞%活性氧%二甲基精氨酸二甲胺水解酶%非对称性二甲基精氨酸
高糖%衰老細胞%活性氧%二甲基精氨痠二甲胺水解酶%非對稱性二甲基精氨痠
고당%쇠로세포%활성양%이갑기정안산이갑알수해매%비대칭성이갑기정안산
High glucose%Cell senescence%Reactive oxygen species%Dimethylarginine dimethylaminohydrolase%Asymmetric dimethylarginine
目的 观察高糖加速内皮细胞衰老过程中细胞内活性氧(ROS)水平与二甲基精氨酸二甲胺水解酶-非对称性二甲基精氨酸(DDAH-ADMA)系统的变化.方法 正常糖浓度培养液(5.5 mmol/L)和高糖培养液(11.0、22.0、33.0 mmol/L)作用于人脐静脉内皮细胞48 h后,用β-半乳糖苷酶染色鉴定衰老细胞,聚合酶链反应-酶联免疫吸附法(PCR-ELISA)检测端粒酶活性,流式细胞仪检测细胞内ROS水平,液质联用仪检测细胞上清液中ADMA含量及DDAH活性.结果 与正常糖浓度组相比,11.0、22.0、33.0 mmol/L高糖浓度组β-半乳糖苷酶染色阳性细胞率明显增高[(7.00±1.73)%、(12.67±2.03)%、(16.00±2.26)%比(4.00±1.33)%,P＞0.05、P＜0.05、P＜0.05],端粒酶活性显著下降[(91.32±4.01)%、(78.44±3.78)%、(56.04±3.35)%比100%,均P＜0.05];随细胞衰老程度加重,细胞内ROS水平(mfi)显著升高(159.84±27.52、188.99±18.77、244.56±20.96比117.11±18.76,P＜0.05或P＜0.01),ADMA水平(μmol/L)显著升高(0.78±0.14、0.88±0.18、1.08±0.15比0.70±0.12,P＞0.05、P＜0.05、P＜0.05),DDAH活性显著下降[(91.32±4.01)%、(78.44±3.78)%、(56.04±3.35)%比100%,均P＜0.05].结论 高糖可加速内皮细胞衰老进程,其机制可能与氧化应激水平增强、抑制DDAH活性使ADMA含量增加有关.
目的 觀察高糖加速內皮細胞衰老過程中細胞內活性氧(ROS)水平與二甲基精氨痠二甲胺水解酶-非對稱性二甲基精氨痠(DDAH-ADMA)繫統的變化.方法 正常糖濃度培養液(5.5 mmol/L)和高糖培養液(11.0、22.0、33.0 mmol/L)作用于人臍靜脈內皮細胞48 h後,用β-半乳糖苷酶染色鑒定衰老細胞,聚閤酶鏈反應-酶聯免疫吸附法(PCR-ELISA)檢測耑粒酶活性,流式細胞儀檢測細胞內ROS水平,液質聯用儀檢測細胞上清液中ADMA含量及DDAH活性.結果 與正常糖濃度組相比,11.0、22.0、33.0 mmol/L高糖濃度組β-半乳糖苷酶染色暘性細胞率明顯增高[(7.00±1.73)%、(12.67±2.03)%、(16.00±2.26)%比(4.00±1.33)%,P＞0.05、P＜0.05、P＜0.05],耑粒酶活性顯著下降[(91.32±4.01)%、(78.44±3.78)%、(56.04±3.35)%比100%,均P＜0.05];隨細胞衰老程度加重,細胞內ROS水平(mfi)顯著升高(159.84±27.52、188.99±18.77、244.56±20.96比117.11±18.76,P＜0.05或P＜0.01),ADMA水平(μmol/L)顯著升高(0.78±0.14、0.88±0.18、1.08±0.15比0.70±0.12,P＞0.05、P＜0.05、P＜0.05),DDAH活性顯著下降[(91.32±4.01)%、(78.44±3.78)%、(56.04±3.35)%比100%,均P＜0.05].結論 高糖可加速內皮細胞衰老進程,其機製可能與氧化應激水平增彊、抑製DDAH活性使ADMA含量增加有關.
목적 관찰고당가속내피세포쇠로과정중세포내활성양(ROS)수평여이갑기정안산이갑알수해매-비대칭성이갑기정안산(DDAH-ADMA)계통적변화.방법 정상당농도배양액(5.5 mmol/L)화고당배양액(11.0、22.0、33.0 mmol/L)작용우인제정맥내피세포48 h후,용β-반유당감매염색감정쇠로세포,취합매련반응-매련면역흡부법(PCR-ELISA)검측단립매활성,류식세포의검측세포내ROS수평,액질련용의검측세포상청액중ADMA함량급DDAH활성.결과 여정상당농도조상비,11.0、22.0、33.0 mmol/L고당농도조β-반유당감매염색양성세포솔명현증고[(7.00±1.73)%、(12.67±2.03)%、(16.00±2.26)%비(4.00±1.33)%,P＞0.05、P＜0.05、P＜0.05],단립매활성현저하강[(91.32±4.01)%、(78.44±3.78)%、(56.04±3.35)%비100%,균P＜0.05];수세포쇠로정도가중,세포내ROS수평(mfi)현저승고(159.84±27.52、188.99±18.77、244.56±20.96비117.11±18.76,P＜0.05혹P＜0.01),ADMA수평(μmol/L)현저승고(0.78±0.14、0.88±0.18、1.08±0.15비0.70±0.12,P＞0.05、P＜0.05、P＜0.05),DDAH활성현저하강[(91.32±4.01)%、(78.44±3.78)%、(56.04±3.35)%비100%,균P＜0.05].결론 고당가가속내피세포쇠로진정,기궤제가능여양화응격수평증강、억제DDAH활성사ADMA함량증가유관.
Objective To investigate the changes in reactive oxygen species (ROS) and dimethylarginine dimethylaminohydrolase-asymmetric dimethylarginine (DDAH-ADMA) system in the process of endothelial cell senescence after exposure to high glucose. Methods The human umbilical vein endothelial cells (HUVECs) were cultured with different concentrations of glucose, e.g. 5. 5 mmol/L (normal level),and high levels as 11. 0, 22. 0 and 33. 0 mmol/L. for 48 hours, respectively. Subsequently, SA-β-gal staining was used to evaluate senescence of cells. Telomerase activity was detected by polymerase chain reactionenzyme linked immunosorbent assay (PCR-ELISA). The intracellular ROS level was measured by flow cytometry. The ADMA concentration and DDAH activity were determined with high-performance liquid chromatography. Results Compared with normal glucose concentration group, after the endothelial cells were treated with high glucose concentration (11. 0 - 33. 0 mmol/L) for 48 hours, the number of SA-β-gal positive cells was increased significantly [(7.00±1. 73)%, (12. 67±2. 03)%, (16. 00±2. 26)% vs. (4. 00±1.33)%, P＞0.05, P＜0.05, P＜0.05] and the telomerase activity was inhibited dramatically [(91. 32±4.01)%, (78. 44±3. 78)%, (56. 04±3. 35)% vs. 100%, all P＜0. 05]. The ROS level (mfi) was increased in all high glucose groups (159. 84±27. 52, 188. 99±18. 77, 244. 56±20. 96 vs. 117.11±18. 76, P＜0. 05 or P＜0. 01). At the same time, the ADMA (μmol/L) production was increased (0. 78±0. 14, 0. 88±0.18,1. 08±0.15 vs. 0. 70±0. 12, P＞0. 05, P＜0. 05, P＜0. 05), and DDAH activity was decreased [(91. 32±4.01)%, (78.44±3.78)%, (56. 04± 3. 35)% vs. 100%, all P＜0.05]. Conclusion High glucose can accelerate endothelial cells senescence in dose-dependent manner and the underlying mechanism may be related to an increased oxidative stress and change in DDAH-ADMA system.
